The proposed will elucidate regulatory controls in the replication of mitochondrial DNAs. Replicative forms will be isolated from thymidine kinase-minus lines of doubly cloned mouse and human cells. A displacement loop model for human mitochondrial DNA replication will be established and compared to that previously constructed for mouse mitochondrial DNA. Control of replication at the level of transcription will be examined through the use of dark-field electron microscopy of transcription complexes and by density gradient and velocity centrifugation of pulse and pulse-chase labeled components. Regulation mediated through mitochondrial specific DNA nucleases in displacement replication will be detailed. Altered replicative forms produced in the stimulation of mitochondrial DNA synthesis by infection of sensitive cells with oncogenic viruses will be examined. The sites on mouse and human mitochondrial DNA where regulatory elements function will be determined relative to known mitochondrial genes and secondary structures by techniques of electron microscopy, gel electrophoresis and thermal denaturation of restriction endonuclease and Pseudomonas endonuclease fragments. Parallel studies will be conducted with circular oligomer and catenated forms of mitochondrial DNA purified from human leukemic white cells before and after patients are treated with antileukemic drugs. Examination of replicative forms will permit construction of a model for displacement replication of human circular dimer mitochondrial DNA. Intramitochondrial distributions of catenated monomeric and oligomeric forms may reflect spatial disposition and relative activities of DNA nucleases in leukemic cell mitochondria and will be assessed by electron microscopy of topological configurations differentially contrasted by vacuum deposition of heavy metals. Altered mitochondrial ultrastructures in human leukemic myeloblasts will be correlated with frequencies of complex mitochondrial DNAs.